Hello!
I have got a problem. I did sequencing of Internal Transcribed Spacer (ITS) using two primers ITS1 and ITS4. I used the same forward and reverse primers for sequencing reaction. I anticipated to obtain two sequnces (forward and reverse). But when I used the sequncher software to make the consensus sequence, the two sequences have the same polarity. I retried but always got the same result. Basically the sequences obtained from forward and reverse primers should be in opposite direction. I can not understand why it always happened with ITS region. I have no problem with cpDNA sequencing and get the sequences in opposite direction with cpDNA sequencing. PLEASE HELP. Thank you in advance!!!!!!
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Adrian K
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Posted 19 November 2009 - 09:59 PM
aashraya, on Nov 15 2009, 06:43 PM, said:
Hello!
I have got a problem. I did sequencing of Internal Transcribed Spacer (ITS) using two primers ITS1 and ITS4. I used the same forward and reverse primers for sequencing reaction. I anticipated to obtain two sequnces (forward and reverse). But when I used the sequncher software to make the consensus sequence, the two sequences have the same polarity. I retried but always got the same result. Basically the sequences obtained from forward and reverse primers should be in opposite direction. I can not understand why it always happened with ITS region. I have no problem with cpDNA sequencing and get the sequences in opposite direction with cpDNA sequencing. PLEASE HELP. Thank you in advance!!!!!!
I have got a problem. I did sequencing of Internal Transcribed Spacer (ITS) using two primers ITS1 and ITS4. I used the same forward and reverse primers for sequencing reaction. I anticipated to obtain two sequnces (forward and reverse). But when I used the sequncher software to make the consensus sequence, the two sequences have the same polarity. I retried but always got the same result. Basically the sequences obtained from forward and reverse primers should be in opposite direction. I can not understand why it always happened with ITS region. I have no problem with cpDNA sequencing and get the sequences in opposite direction with cpDNA sequencing. PLEASE HELP. Thank you in advance!!!!!!
You mean consensus or contiq? as far as I concern, you should do a contiq and then get the consensus... correct me if i'm wrong.
Expecting the world to treat you fairly because you are a good person is like expecting the lion not to attack you because you are a vegetarian.
..."best of our knowledge, as far as we know this had never been reported before, though I can't possible read all the published journals on earth, but by perform thorough search in google, the keywords did not match any documents"...
"what doesn't kill you, makes you stronger"---Goddess Casandra reminds me to be strong
"It's all just DNA. Do it."---phage434
..."best of our knowledge, as far as we know this had never been reported before, though I can't possible read all the published journals on earth, but by perform thorough search in google, the keywords did not match any documents"...
"what doesn't kill you, makes you stronger"---Goddess Casandra reminds me to be strong
"It's all just DNA. Do it."---phage434
